Since 1990 we have performed over 275 simultaneous pancreas and kidney (SPK) transplants in patients with type 1 diabetes (T1D) and end-stage renal disease. We are presently following 225 recipients. While all patients became insulin-independent, 34 (15%) have returned to hyperglycemia after a mean follow-up of 5.7 years. Our preliminary data demonstrate the presence of diabetes-associated autoantibodies preceding the return of hyperglycemia in approximately 65% of the patients with hyperglycemia and in 40% of normoglycemic SPK recipients. The observed frequency of autoimmunity in association with increased risk of hyperglycemia is higher than previously reported in smaller and earlier studies. Patients with selective loss of insulin secretion and without evidence of rejection were studied in detail and were found to have the cardinal features of recurrence of autoimmunity, including autoantibodies, insulitis and the presence of autoreactive T cells in peripheral blood assessed by recently developed tetramer-based assays. Thus, our data provide evidence that recurrent autoimmunity is a clinical problem of great significance that was previously underestimated. We have assembled a team of investigators with experience in pancreas transplantation (Burke, Miami) and the immunology of T1D (Pugliese, Miami;Nepom, Seattle) to study the key immunological events associated with recurrence of autoimmunity in SPK recipients. We plan to (1) retrospectively and prospectively analyze our cohort of 225 SPK patients to determine the frequency and time course of autoantibody recurrence and the predictive value of autoantibodies for recurrence of disease;(2) prospectively follow pancreas transplant recipients and assess humoral and cellular responses on follow-up samples by: (a) monitoring autoantibody levels, (b) monitoring and phenotyping autoreactive T cells in peripheral blood, (c) assessing islet/pancreas pathology from biopsies performed in recipients with consistent recurrence of multiple autoantibodies and (d) monitoring and phenotyping autoreactive T cells from the pancreatic infiltrate obtained by pancreatic transplant explants and/or biopsies. The proposed studies will define the relationship between autoantibodies, autoreactive T cells, insulitis and insulin secretion in SPK recipients. Methodological advances such as tetramer-based assays offer an unprecedented opportunity to characterize epitope specificity, avidity and the functional status of autoreactive T cells and compare these parameters in cells from peripheral blood and the pancreatic infiltrate. These studies will provide critical information about the immunological mechanisms regulating recurrence of autoimmunity in SPK recipients and assess the predictive value of laboratory tests that may find clinical application in the pancreas transplant setting. The knowledge gained should also be relevant to islet cell transplantation and spontaneous disease.